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Description
Bst DNA Polymerase, Large fragmentProduct Specification Synonyms DNA polymerase I Expression System E. coli Molecular Weight 67kDa (Reducing) Purity 95% by SDS PAGE Tags & Cleavage sites Tag His Tag Storage Buffer 50 mM KCl10 mM Tris HCl0. 1 mM EDTA1 mM DTT0. 1% Triton X 10050% GlycerolpH 7. 1 @ 25C Stability & Storage Store at 25 ~ 15 for 2 years Reference 1. Nucleic Acids Research, 2000, 28(12): E63. 2. Chemical Communications, 2014, 50(28): 3747 3749. 3. Current Protocols in
Product Specification
| Synonyms | DNA polymerase I |
| Expression System | E.coli |
| Molecular Weight | 67kDa (Reducing) |
| Purity | ＞95% by SDS-PAGE |
| Tags & Cleavage sites | / |
| Tag | His Tag |
| Storage Buffer | 50 mM KCl、10 mM Tris-HCl、0.1 mM EDTA、1 mM DTT、0.1% Triton® X-100、50% Glycerol、pH 7.1 @ 25°C |
| Stability & Storage | Store at -25 ~ -15℃ for 2 years |
| Reference | 1. Nucleic Acids Research, 2000, 28(12):E63. |
Background
Bst DNA Polymerase, Large Fragment is the portion of the Bacillus stearothermophilus DNA Polymerase protein that contains the
5´ → 3´ polymerase activity, but lacks 5´ →3´ exonuclease activity. It can be use to isothermal amplification (LAMP), DNA sequencing through high GC regions and Rapid Sequencing from nanogram amounts of DNA template.
Components
50 mM KCl、10 mM Tris-HCl、0.1 mM EDTA、1 mM DTT、0.1% Triton® X-100、50% Glycerol、pH 7.1 @ 25°C
10* Reaction Buffer: 200 mM Tris-HCl、100 mM (NH4)2SO4、20mM MgSO4、100 mM KCl、1% Tween® 20、pH 8.8@25°C
Magnesium Sulfate (MgSO4) Solution：100mM MgSO4
Protocol
Incubate the following reaction at 65°C for 30–60 minutes
Component |
Final Concentration |
10X Isothermal Amplification Buffer II |
1X (contains 2 mM MgSO4) |
MgSO4 (100 mM) |
6 mM (8 mM total) |
dNTP Mix (10 mM) |
1.4 mM each |
FIP/BIP Primers (25X) |
1.6 µM |
F3/B3 Primers (25X) |
0.2 µM |
LoopF/B Primers (25X) |
0.4 µM |
Bst DNA Polymerase (8,000 U/ml) |
320 U/ml |
DNA or RNA Sample |
> 10 copies or more |
Nuclease-free Water |
to 25 µl |
Total Reaction Volume |
25 µl |
Guidelines
1.Bst DNA Polymerase does not exhibit 3´→ 5´ exonuclease activity.
2.100 µg/ml BSA or 0.1%Triton X-100 is required for long term storage.
3.Reaction temperatures above 70°C are not recommended.
4.Bst DNA Polymerase, Large Fragment cannot be used for thermal cycle sequencing or PCR
Unit Definition
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